Related Experiment Video
Updated: Jul 24, 2025
Multi-target Parallel Processing Approach for Gene-to-structure Determination of the Influenza Polymerase PB2 Subunit
Published on: June 28, 2013
Advances in the expression and purification of human PARP1: A user-friendly protocol
Carlota J F Conceição1, Bruno A Salgueiro2, Paulo A Ribeiro3
1CEFITEC, Department of Physics, NOVA School of Science and Technology, Universidade NOVA de Lisboa, 2829-516, Caparica, Portugal; Laboratory of Instrumentation, Biomedical Engineering and Radiation Physics (LIBPhys-UNL), Department of Physics, NOVA School of Science and Technology, Universidade NOVA de Lisboa, 2829-516, Caparica, Portugal.
Abstract:
The PARP1 (Poly (ADP-ribose) polymerase 1) enzyme is essential for single and double-strand break repair in humans. Alterations affecting PARP1 activity have severe consequences for human health and are associated with pathologies like cancer, and metabolic and neurodegenerative disorders. Here, we have developed a fast and easy procedure for the expression and purification of PARP1. Biologically active protein was purified to an apparent purity > 95%, with only two purification steps. A thermostability analysis revealed that PARP1 possessed improved stability in 50 mM Tris-HCl pH 8.0 (Tm = 44.2 ± 0.3 °C), thus this buffer was used throughout the whole purification procedure. The protein was shown to bind to DNA and has no inhibitor molecules bound to the active site. Finally, the yield of the purified PARP1 protein is sufficient for both biochemical, biophysical and structural analysis. The new protocol provides a fast and simple purification procedure while producing similar protein quantities to what has been described previously.
